Serdar Bulun, MD

George H. Gardner Professor of Clinical Gynecology

Chief, Division of Reproductive Biology ResearchDepartment of Obstetrics and Gynecology

Northwestern University, Chicago, IL

Pathogenesis of Pain in Endometriosis

ENDOMETRIOSIS• Common cause of chronic pelvic pain and infertility

• Polygenic inheritance and multifactorial etiology

• 3-6 million women of reproductive age are affected in the USA

• Responsible for 10%-15% of all hysterectomies performed in the USA

GLANDS

STROMA

MEDICAL TREATMENT OPTIONS

• Oral contraceptives

• GnRH analogs

• Progestins

• Aromatase inhibitors

• Selective Progesterone Receptor Modulators

ANDROSTENEDIONE

aromatase

adrenaladrenal

OVARYOVARYANDROSTENEDIONE

GnRH-aOC

aromatase inhibitor

FAT AND SKIN

ANDROSTENEDIONE ESTRONE

3-Estradiol

1-Estradiol

2-Estradiol

CHOLESTEROL

StAR

ENDOMETRIOSIS

aromatase

Endometrium fromDisease-free Women

Endometrium fromWomen with Endometriosis

Endometriosis

1

2

3

PRECURSOR

E

E

PGE2

PGE2

PGE2

PRECURSOR aromataseESTROGEN

PRECURSOR aromatase ESTROGEN

COX-2PGE2PRECURSOR

COX-2 PGE2PRECURSOR

COX-2 PGE2PRECURSOR

Noble, et al, JCEM, 1996; Ota, et al, 2001

SF1

SF1

CYTOPLASM

Src

DNATarget geneNRE

PROTEIN PRODUCTSOF TARGET GENES

BIOLOGIC FUNCTIONe.g., DIFFERENTIATION

NUCLEUS

NR

NRP

membranereceptor

MAPK

TF

classical

nonclassical

NRP

NRP

steroidreceptor

NUCLEAR RECEPTORS (NR)

NRP

orphan nuclear receptor

fold

dif

fere

nce

(m

RN

A b

y re

al-t

ime

PC

R)

ERαERβ TotalPR

PR-B

ENDOMETRIUM (TISSUE)

OVARIAN ENDOMETRIOMA (TISSUE)

1

2

3

4

5

6

7

8

9

108.9

142.4 9.3

14.1

Xu Q et al, BOR 2007

SF

1 m

RN

A

0.2

0.4

0.6

0.8

1.0

E-IUM E-OSIS

>12,000

NUCLEAR RECEPTORS AND ENDOMETRIOSIS

EPIGENETIC DEFECTS IN ENDOMETRIOSIS

SF-1 HYPOMETHYLATION

Methylation of CpG islands at promoters and gene expression

Unmethylated CpG site

Methylated CpG site

exon1 exon2 exon3

exon1 exon2 exon3

on

off

Promoter CpG island

SF1 GENE PROMOTER

NORMAL SUBJECTS

PATIENTS WITHENDOMETRIOSIS

exon1 exon2

exon1 exon2

Transcription coactivatingcomplex

HDAC1

exon1 exon2

exon1 exon2

exon1 exon2

exon1 exon2

SF1on

offMeCP2SF1

ENDOMETRIOSIS

ENDOMETRIUM

Xu Q et al, JCEM 2007

mSin3A

SF1 PROMOTER IS NOT METHYLATED IN ENDOMETRIOSIS

unmethylated CpG island

methylated CpG island

ROLE OF SF-1 IN

ESTROGEN

PRODUCTION

1

2

3

4

G

E-IUM E-OSIS

LRH

-1 m

RN

A (

amol

/mg

18S

)

0.5

1

1.5

2.0

E

E-IUM E-OSIS

P45

0aro

m m

RN

A (

amol

/mg

18S

)

*

un-detectable

FS

F-1

mR

NA

(am

ol/m

g 18

S)

0.2

0.4

0.6

0.8

1.0

E-IUM E-OSIS

*

StA

R m

RN

A

(am

ol/m

g 18

S)

20

40

60

80

100

120

140

160

E-IUM E-OSIS

A*p<0.01

P45

0scc

mR

NA

(am

ol/m

g 18

S)

0.5

1

1.5

2.0

2.5

B

E-IUM E-OSIS

*p<0.001

E-IUM E-OSIS

3HS

DB

II (

amol

/mg

18S

)

0.1

0.2

0.3

0.4

0.5

0.6

0.7

C

*p<0.01

p<0.001 p<0.001 p<0.05

*

E-IUM E-OSIS

P45

0c17

mR

NA

(am

ol/m

g 18

S)

0.05

0.1

0.15

0.2

0.25

0.3

0.35

D

*p<0.05

Attar and Tokunaga et al, JCEM, 2009

PERITONEAL ENDOMETRIOSIS

0

5

10

15

20

25

30

StA

R (

amol

/mg

18S

)H

SD

3BI (

amol

/mg

18S

)

HS

D3B

II (a

mol

/mg

18S

)

*

*

*

*

*

*p= 0.032

PGE2: - + - +E-IUM E-IUM E-OSIS E-OSIS

0.00

0.01

0.02

0.03

0.04

0.05

0.06

0.00

0.05

0.10

0.15

0.20

0.25

PGE2: - + - +E-IUM E-IUM E-OSIS E-OSIS

PGE2: - + - +

E-IUM E-IUM E-OSIS E-OSIS

*

E-IUM E-IUM E-OSIS E-OSIS

PGE2: - + - +

P45

0aro

m (

amol

/mg

18S

)

0.0

0.2

0.4

0.6

0.8

1.0

1.2 *

*

*p= 0.030

*p= 0.05 *p= 0.012

E-IUM E-IUM

PGE2: - + - +

0.00

0.02

0.04

0.06

0.08

0.10

0.12

P45

0c17

(am

ol/m

g 18

S)

*

*

E-OSIS E-OSIS

*p= 0.042

*

*

E-IUM E-IUM

P45

0scc

(am

ol/m

g 18

S)

0

2

4

6

8

10

12

14

PGE2: - + - +E-OSIS E-OSIS

*p= 0.048

A B

C D

E F

OVARIAN ENDOMETRIOTIC STROMAL CELLS

Attar and Tokunaga et al, JCEM, 2009

HO

O

O

O

OH

HO

O

HO

C

CH3

O

OH

HO

O

O

C

CH3

O

O

C

CH3

OOH

HO

C

CH3

O

HO

OH

cholesterol

MITOCHONDRION

pregnenolone

17-hydroxypregnenolone

dehydroepiandrosterone

StAR

P450scc

P450c17

P450c17

androstenedione testosterone

estrone ESTRADIOL

17-hydroxyprogesterone

PROGESTERONE

P450c17

P450c17

P450arom P450arom

HSD3B2

HSD3B2

HSD3B2 HSD17B1

HSD17B1

6

12

34

5

9

7

810

1112

13

14 1516

17

18

19

21 2022

23

252427

26

++++++++++

ENDOMETRIOSIS

nmol/L

pmol/L

estroneandrostenedione

estradiol

aromatase

PGE2COX-2

++adrenal

AA

++

ENDOMETRIOSIS TISSUE

aromatase inhibitor

CHOLESTEROL

++

Attar and Tokunaga, et al, JCEM, 2009Takayama et al, Fertil Steril, 1998Ailawadi et al, Fertil Steril, 2004Amsterdam et al, Fertil Steril, 2005

Noble et al, JCEM, 1997Zeitoun et al, Mol Endo, 1999Tamura et al, JBC, 2002Tamura et al, JCEM, 2002Tamura et al, JCEM, 2002

IL-1VEGF

++SF1

StAR

scc

3-HSD-2

17-OH-lase

17/20-lyase

Hypothalamus

Pituitary FSHLH

Postmenopausal on AI

Ovary

Peripheral Tissues

Endometriosis

Aromatase

PeripheralAromatase

No Follicular

Aromatase

AI

Aromatase

Takayama et al, Fertil Steril, 1998; Ailawadi et al, Fertil Steril, 2004; Soysal et al, Human Reproduction, 2004; Amsterdam et al, Fertil Steril, 2005; Attar and Bulun Fertil Steril, 2006

Follicle Development

Prememopausal on AI(Hypothetical)

Aromatase

FSHLH

AromataseE2

Aromatase

AI

PeripheralAromatase

Prememopausal on AI+ OC or P

Aromatase

Aromatase

PeripheralAromatase

AI

OC P

GnRHa

AromataseE2

FSHLH

Pre- and Post-treatment Pain Scores (n=10)

0

5

10

0

5

10

Ave

rage

Pai

n S

core

Baseline post-treatmentp=0.0028

PREMENOPAUSAL WOMEN TREATED WITH LETROZOLE AND NORETHINDRONE ACETATE

Ailawadi, et al, Fertil Steril, 2004

Before Treatment

PREMENOPAUSAL WOMEN TREATED WITH LETROZOLE AND NORETHINDRONE ACETATE

Ailawadi, et al, Fertil Steril, 2004

Before Treatment

PREMENOPAUSAL WOMEN TREATED WITH LETROZOLE AND NORETHINDRONE ACETATE

Ailawadi, et al, Fertil Steril, 2004

After Treatment for 6 Months

PREMENOPAUSAL WOMEN TREATED WITH LETROZOLE AND NORETHINDRONE ACETATE

Ailawadi, et al, Fertil Steril, 2004

Mild hot flushes (2/day – 1/week)

Sleepiness

Mood swings

Decreased libido

No bone loss

SIDE EFFECTS

TREATMENT RECOMMENDATIONS• An aromatase inhibitor is the treatment of choice for persistent

postmenopausal endometriosis

• A combination of an aromatase inhibitor with an OC or GnRH analog may be used to treat endometriosis-related pain not responding to conventional treatments

• Await results of randomized studies for the routine use of aromatase inhibitors for premenopausal endometriosis, uterine leiomyomata and ovulation induction

EPIGENETIC DEFECTS IN ENDOMETRIOSIS

ER HYPOMETHYLATION

Bisulfite sequencing: ERβ promoter

Exon 0N

Exon 0N

TSS +1

100bpBisulfite sequence

Classical CpG island 556bp

Exon 1

-163 -158 -133 -131 -129 -126 -123 -119 -107 -104 -90 -74 -48 -163 -158 -133 -131 -129 -126 -123 -119 -107 -104 -90 -74 -48

1

2

3

4

5

6

7

8

1

2

3

4

5

6

7

8

-163 -158 -133 -131 -129 -126 -123 -119 -107 -104 -90 -74 -48 -163 -158 -133 -131 -129 -126 -123 -119 -107 -104 -90 -74 -48

1

2

3

4

5

6

7

8

1

2

3

4

5

6

7

8

44481bp

Exon 0K

10957bp

NORMAL SUBJECTS

PATIENTS WITHENDOMETRIOSIS

P< 0.001

ERgene

MeCP2

HDAC1mSin3a

Pr 0N

CpG island

Pr 0K

Methylated

Pr 0N

CpG island

Pr 0K

Unmethylated

+CBP

ENDOMETRIUM ENDOMETRIOSIS

ROLE OF ER IN

PROSTAGLANDIN

PRODUCTION

PROSTAGLANDIN AND ESTRADIOLBIOSYNTHETIC PATHWAYS

ANDROSTENEDIONE

ESTRONE

ESTRADIOL

HSD17B1

Aromatase

PGE2

PGH2

ARACHIDONIC ACID

COX-2

mPGES-1

PHOSPHOLIPIDSPLA2

?

PROSTAGLANDIN AND ESTRADIOLBIOSYNTHETIC PATHWAYS

ANDROSTENEDIONE

ESTRONE

ESTRADIOL

HSD17B1

Aromatase

PGE2

PGH2

ARACHIDONIC ACID

COX-2

mPGES-1

PHOSPHOLIPIDSPLA2

ER

IL-1VEGF

ROLE OF ER IN

PROGESTERONE

RESISTANCE

MensesProliferativeendometrium

E2

Secretoryendometrium

E2 + P

17-HSD-2: _ ++++_

(Epithelial)

E2 E1:_ ++++

_

Bulun SE, et al, MCE, 2006

HO

OH

HO

O

HSD17B2

PROGESTERONE

HO

OH

HSD17B2

+

PROGESTERONE

progesterone resistance

EUTOPICENDOMETRIUM

ENDOMETRIOSIS

E1

E2

E1E2

HO

O

Zeitoun, et al, JCEM, 1998Yang, et al, Mol Endo, 2001

T47

D

Cycle Day: 3 6 10 12 14 16 18 24 26

PR-B

PR-A

EUTOPICENDOMETRIUM

PR-B

PR-AENDOMETRIOSIS

Attia, et al, JCEM, 2000

ERgene

Prom 0N

CpG island

ER

PRgene

PR-APR-B

ER

+

ER

ENDOMETRIOSISMeCP2

HDAC1mSin3a

ER

ER

PR-BPR-A

ER ER ER

ERgene

Prom C Prom A

Trukhacheva et al, JCEM, 2009

Bulun, SE, NEJM 2009Cheng, Y, JCEM, 2008

BIG PICTURE

PGE2

estradiolCHOLESTEROL

SF1COX-2

ER ER

PR

retinoic acid

estroneHSD17B2 arom

methylation

methylation

Bulun et al, NEJM, 2009

Bulun, SE, NEJM 2009

Menses

ERSF1

Painful menses

OTHERS

SURVIVAL AND IMPLANTATION OF ENDOMETRIOTIC CELLS

FAILURE OF CELLS TO IMPLANT

peritoneum peritoneum

NORMAL ENDOMETRIOSIS

methylatedpromoter

unmethylatedpromoter

CELLS CLEARED BY IMMUNE SYSTEM

+

ROLE OF EPIGENETICS INMECHANISM OF

ENDOMETRIOSIS

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